The present invention relates to a novel F0F1-ATPase, a DNA encoding the F0F1-ATPase, a method for producing the F0F1-ATPase, and a method for producing nucleoside 5′-triphosphate; using the F0F1-ATPase.
F0F1-ATPase plays principal roles in the biological energy metabolism, because the enzyme has an activity of generating adenosine 5′-triphosphate (ATP) as an energy source of organisms, by utilizing the gradient of proton concentration between the intramembrane and the extramembrane. Therefore, utilization of F0F1-ATPase enables us to develop a living thing with improved energy metabolism.
Herein, F0F1-ATPase is synonymous with H+-ATPase.
Because it has been known that the activity of F0F1-ATPase varies, depending on the change of outer environment, such as pH, the utilization of F0F1-ATPase can provide a living thing adaptable to the change of outer environment [Mol. Microbiol., 33, 1152 (1999), J. Bacteriol., 176, 5167 (1994)].
F0F1-ATPase is a protein complex comprising a soluble catalytic sector F1 and a transmembrane sector F0 functioning as proton channel. In organisms such as Escherichia coli and Bacillus subtilis, F1 is composed of five subunits of α, β, γ, δand ε, while F0 is composed of three subunits of a, b and c [Annu. Rev. Biochem., 66, 717 (1997)].
Concerning the F0F1-ATPase gene, the gene was isolated from Escherichia coli [Biochem. J., 224, 799 (1984)], Bacillus subtilis [J. Bacteriol., 176, 6802 (1994)], Bacillus megaterium [J. Biol. Chem., 264, 1528 (1989)], Bacillus firmus [Mol. Gen. Genet., 229, 292 (1991)], Bacillus sp. PS3 [Biochim. Biophys. Acta,. 933, 141 (1988)], Methanosarcina barkeri [Biochem. Biophys. Res. Commun., 241, 427 (1997)], Lactobacillus acidophilus [Mol. Microbiol., 33, 1152 (1999)], Rhodobacter capsulatus [J. Bacteriol., 180, 416 (1998), Arch. Microbiol., 170, 385 (1998)] and the like, but no gene derived from microorganisms belonging to the genus Corynebacterium has been isolated yet.
Regarding the production of nucleoside 5′-triphosphate, methods using microorganisms (Japanese Published Unexamined Patent Application No.107593/1979; Japanese Published Unexamined Patent Application No. 51799/1984; J. Ferment. Bioeng., 68, 417 (1989)) and a method using enzymes (WO 98/22614) have been known. However, the productivity of nucleoside 5′-triphosphate is insufficient.